title: Purification of a rat liver cytosolic sulfotransferase responsible for the conjugation of digitoxigenin.
abstract: Previous investigations on the digitoxin metabolism hardly considered the role of the sulfate ester conjugation. Therefore, this study examined whether digitoxin (dt-3) or one of its cleavage products might be sulfated in vitro. It was proven that digitoxigenin (dt-0) is by far the best substrate for the cytosolic sulfotransferases (ST). Digitoxigenin-monodigitoxoside (dt-1) and digitoxigenin-bisdigitoxoside (dt-2) are sulfated in trace amounts whereas dt-3 is not sulfated at all. The purification of the responsible enzyme was performed by liquid chromatography on Q-Sepharose and hydroxyapatite. During the purification procedure this enzymatic activity corresponded exactly to that towards dehydroepiandrosterone (DHEA). The 134-fold purified and gel electrophoretically homogeneous enzyme protein (Mr 33,000) showed a Vmax of 12.5 nmoles dt-0 sulfate/min mg protein and a KM of 37 mumol/L. The purified enzyme conjugated dt-1 and dt-2 in trace amounts only and was inhibited competitively by DHEA. It can be concluded that in the rat a 3 beta-hydroxy-steroid sulfotransferase is responsible for the sulfation of dt-0. The purified enzyme reacts with dt-1, dt-2 and digoxigenin (dg-0) in traces only, a sulfation of dt-3 is not detectable.
authors.affiliation: Institute for Legal Medicine, University of Hamburg, Federal Republic of Germany.
authors.forename: A
authors.initials: A
authors.lastname: Johannes
authors.forename: L
authors.initials: L
authors.lastname: von Meyerinck
authors.forename: A
authors.initials: A
authors.lastname: Schmoldt
mesh_terms: D000818:Animals; D002852:Chromatography, Ion Exchange; D003600:Cytosol / Q000201:enzymology; D004073:Digitoxigenin / Q000378:metabolism*; D004074:Digitoxin / Q000378:metabolism*; D004076:Digoxigenin / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D005260:Female; D007700:Kinetics; D008099:Liver / Q000201:enzymology*; D051381:Rats; D011919:Rats, Inbred Strains; D015238:Sulfotransferases / Q000302:isolation & purification*; D013466:Sulfurtransferases / Q000378:metabolism
chemical_list: D004073:Digitoxigenin; D004074:Digitoxin; D013466:Sulfurtransferases; D015238:Sulfotransferases; C028152:alcohol sulfotransferase; D004076:Digoxigenin